Identification and quantification of anabolic steroids in imported frozen beef muscles in sulaimani market using HPLC

Abstract

In Kurdistan region of Iraq, little information about steroid hormone residues in animal products is available. The work was developed for determination of eight natural and synthetic residues of steroid hormones in five trademarks (Kilcoy,Veal Topsides, Pure south, P.Van Horten & Zn, and Creek stone Farms) of beef muscles imported from five countries. The determined values were compared with the national limits to conclude either is in legal values. The imported frozen beef muscle samples (n=75) were collected from 10 th January to 10 th of May 2015. Liquid-liquid phase extraction was used, and then amino-propyl cartridges and dual silica used for clean-up. The analyses were quantified by High Performance Liquid Chromatography using a phenyl column coupled to an electrospray ionization UV spectrometer (LC–ESI-UV). The results presented that all meat samples were treated with the eight steroid; whereas, progesterone and estradiol benzoate were in safe level (0.41 ± 0.5 µg/kg and 0.64 ± 0.36 µg/kg respectively) in all meat samples. The rest steroids residues were higher than the national limits except medroxy progesterone which was (0.49 µg/kg) in safe level only in samples of Kilcoy. There was no significant difference among the steroids levels regarding the five trademarks except diethylstilbestrol level which was significantly different (P value = 0.046) between P.Van Horten & Zn and Creek Stone Farms and medroxy progesterone (P value = 0.03) between kilcoy and P.Van Horten & Zn.

Full text

PAK. J. FOOD SCI., 26(2), 2016: 72-82
ISSN: 2226-5899
Pakistan Journal of Food Sciences (2016), Volume 26, Issue 2, Page(s): 72-82
72
Identification and quantification of anabolic steroids in imported
frozen beef muscles in sulaimani market using HPLC
Hazhaow Omar Murad,1* Faraidoon Abdulstar Muhamad,2* Ahmed Yaseen Hamadameen3*
1Department of Microbiology, College of Veterinary Medicine, University of Sulaimani, Sulaimani New, Street
27,Sulaimani City, Kurdistan Region, Northern Iraq
2Department of Surgery and Theriogenology, College of Veterinary Medicine, University of Sulaimani, Sulaimani
New, Street 27, Sulaimani City, Kurdistan Region, Northern Iraq
3Department of Microbiology, College of Veterinary Medicine, University of Sulaimani, Sulaimani New, Street 27,
Sulaimani City, Kurdistan Region, Northern Iraq
Correspondence Author: Ahmad.Hamadamin@univsul.ed.iq
ABSTRACT
In Kurdistan region of Iraq, little information about steroid hormone residues in animal products is available. The
work was developed for determination of eight natural and synthetic residues of steroid hormones in five trademarks
(Kilcoy,Veal Topsides, Pure south, P.Van Horten & Zn, and Creek stone Farms) of beef muscles imported from five
countries. The determined values were compared with the national limits to conclude either is in legal values. The
imported frozen beef muscle samples (n=75) were collected from 10th January to 10th of May 2015. Liquid-liquid
phase extraction was used, and then amino-propyl cartridges and dual silica used for clean-up. The analyses were
quantified by High Performance Liquid Chromatography using a phenyl column coupled to an electrospray ionization
UV spectrometer (LC–ESI-UV). The results presented that all meat samples were treated with the eight steroid;
whereas, progesterone and estradiol benzoate were in safe level (0.41 ± 0.5 µg/kg and 0.64 ± 0.36 µg/kg respectively)
in all meat samples. The rest steroids residues were higher than the national limits except medroxy progesterone which
was (0.49 µg/kg) in safe level only in samples of Kilcoy. There was no significant difference among the steroids levels
regarding the five trademarks except diethylstilbestrol level which was significantly different (P value = 0.046)
between P.Van Horten & Zn and Creek Stone Farms and medroxy progesterone (P value = 0.03) between kilcoy and
P.Van Horten & Zn.
Keywords: Growth Promoting Hormones; Frozen beef muscle, Liquid-Liquid Extraction, HPLC, Sulaimani markets.
INTRODUCTION
Anabolic steroids are a group of synthetic hormones
which could promote the storage of proteins and the
growth of tissues (Kicman, 2008). Indeed, some
hormones are able to create young animals obtain
weight faster and help to decrease the weighting time
and the amount of feed consumed by an animal pre
slaughtering in meat processing area(FDA, 2013). In
the 1970s, anabolic steroids were widely used to
increase growth rate and develop feed conversion
ability animals (Ayyar, 2011). Whereas, further
studies found that they had toxic or carcinogenic
properties; hence, the European Commission (EC) and
the Food and Drugs Administration (FDA) prohibited
their usage (Ricke, 2012). To control not allowed
treatments, it was necessary to establish sensory
analytical methods. Therefore, a many different of
procedures has been described for the determination of
anabolic residue in tissues (Sofos, 2005). Recently, the
EC recommended several methods for the detection of
hormonally active compounds, including high
performance liquid chromatography- (HPLC)
(Corradini and Phillips, 2011) and gas
chromatography-mass spectrometry (GC-MS) (Horie
and Nakazawa, 2000), and immunoassay techniques
such as enzyme linked immunosorbent assay (ELISA)
which has developed for several compounds nowadays
(Peng et al., 2008). The steroid hormones of estrone,
zeranol, diethylstilbestrol, trembolone, medroxy
progesterone, melengestrole acetate, and progesterone
and estradiol benzoate have been licensed as growth

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73
promoters for farm animals in several meat-exporting
countries. By contrast, it has been noticed that high
level of the hormones injected to animals caused to be
higher level of hormone residues and leads several
health issues in humans (Kerth, 2013). By contrast,
many investigation have reported their safety for
animals and consumers, little is known about their fate
after excretion by the animal; as well as, some steroids
remain more than 3 months after implantation (te
Pasetal, 2004). The best standard technique for steroid
analysis has been liquid and gas chromatography yet
(Yan et al., 2009). However, HPLC-UV is a sensitive,
robust and suitable technique for the assay of
hormones (Warriss, 2000), it is necessary to have
analytical methods to extract meat samples used in
HPLC, as well as the complexity to look after
admixture in tissue samples. Hence, it is acceptable
method and has the minimum required performance
limits (MRPLs). Understandably, the required
concentrations and action limits respectively
authorised by CD 96/23/EC, the Community
Reference Laboratories (CRLs) and the National
Reference Laboratory (WIV) (Corradini, 2010). So,
the aim of this work was to apply a sensitive,
applicable and reliable multi-procedure for the
identification and confirmation of eight famous
anabolic steroids in bovine muscle imported in to
Kurdistan region to confirm that how far are they safe
for human consumption.
Materials and Methods
Samples and sampling plan.
The information in (Table 1) elucidate the trade name,
country of origin and company name for various
samples.
Chemicals and Reagents
Anabolic hormone standards and solvents were
provided for extraction and purification steps for
HPLC application. Pure standards of synthetic
hormones used in the study including internal
standards were all purchased from Sigma (Sigma–
Aldrich, Steinhem, Germany). Ethanol (HPLC grade)
and tris (hydroxymethyl) aminomethane were
provided from Merck (Darmstadt, Germany), hexane,
β- glucuronidase, acetonitrile, dichloromethane,
glacial acetic acid and sodium acetate were from
Sigma (Sigma–Aldrich, Steinhem, Germany).
Ultrapure water was produced with a Pure Lab system
(Sation 9000, Spain). In addition, 2%
ammonium/water solution was prepared by adding
8ml ammonium 25% in 92 ml of water. Oasis HLB (60
mg, 3 mL) cartridges were obtained from Waters
(Milford, MA, USA) and Amino Supelclean NH2
cartridges from Supelco (Bellenfonte, IL, USA).
Table 1. Sample collection regarding trade name,
countries origin and companies
Sampling
No.
Trade
Name
Producer
Country
No. of
Sample
Collecting
Period
1
Kilcoy
Australia
3×5
11/1/2015 to
23/5/ 2015
2
Pure
south
New
Zealand
3×5
14/1/2015 to
20/5 2015
3
Veal
Topsid
es
Switzerland
3×5
23/1/2015 to
27/5/2015
4
P.Van
Horten
& Zn
Holand
3×5
15/1/2015 to
27/5/2015
5
Creek
stone
Farms
America
3×5
15/1/2015 to
20/5/2015
* Samples for each company collected from three
different markets and 5 samples in each market for
each type.
The stock standard solutions (1 mg/ml) were prepared
in ethanol and stored at −20oC in the dark. Formic acid
(99%), acetic acid (99%) and trifuoroacetic acid (99%)
were from Acros Organics (New Jersey, USA).
Anhydrous sodium sulphate and ammonium formic
were all in analytical grade (Beijing, China). To avoid
contamination, all the glassware was sterilised for 4 h
at 400oC prior to use. In addition, procedural blanks
were conducted for each batch of samples to ensure
minimal contamination. Spiking and calibration
mixtures at various concentration levels were obtained
by combing aliquots of stock solutions and internal
standards with mobile phase and stored at 4oC.
Instrumentation
HPLC analysis was carried out using a Shimadzu
HPLC system equipped with two LC-20AT Solvent
Delivery Units, a SUS20A gradient controller, and a
SPD-20A UV-VIS Detector (Shimadzu, Kyoto,
Japan). An N-2000 Chromatographic workstation
(Zheda Zhineng Co. Ltd., Hangzhou, China) was used
as a data acquisition system. The analytical column
was purchased from RStech Co., Korea (3 µm particle

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size 50 mm×4.6 mm I.D, C18, 5.0 µm). The mobile
phase was (A) 0.1% Ammonium formate (NH4HCO)
in ultra-pure water. Solvents V/Methanol: Acetonitrile
HPLC grade, and linear gradients0% B to 100%B. The
detection wavelength of the detector was set at 242
nm. And the flow rate 1.4 ml/min at 35oC.
Standard Solution Preparation
According to (Liu et al, 2011), primary stock solutions
were prepared in ethanol at concentration of 1000 ng/
µL−1 and the working solutions were prepared by 100×
and 1000× dilution in ethanol/water (50/50). Solutions
were stored in dark glass container at −20oC. Sodium
acetate buffer 0.2 M (pH 5.4) was prepared by
dissolving 54.5 g sodium acetate in 600 mL 0.2 M
glacial acetic acid. Then, the solution was diluted to 2
L with ultrapure water and the NH2 (100 mg, 1 mL)
cartridges were provided for the purification. Frozen
meat samples (75) were collected from local markets
and saved in fridge temperature. All the samples were
homogenized by a disintegrator separately.
Liquid-Liquid Extraction Samples
To follow (Huopalahti and Henion, 1996), the liquid-
liquid extraction was carried out by using 20g of each
minced meat separately. All samples were
homogenized with 100 ml ethanol. The homogenate
were then vortex-mixed with 10 ml of 0.03M sodium
acetate. The pH of the solution was brought to 4.5 with
glacial acetic acid. The enzymatic hydrolysis was
initiated by adding 100µl of β-glucuronidase. After 8
hours of enzymatic hydrolysis at 37oC, 20 ml of
acetonitrile was added followed by mixing on a vortex
mixer for 30 seconds. The homogenate was then
centrifuged (5000 rpm for 20 min) and the supernatant
(30ml) were then transferred to a clean test tube.
Hexane (8mL) and dichloromethane (2 mL) were
added and mixed by rotation for 3 minutes. Samples
were then centrifuged (2000 rpm for 2 min) and
extracted with 25 mL acetonitrile and transferred to a
20 ml scintillation vial and evaporated to dryness then
re-dissolved in 1 ml of dichloromethane and the
aqueous extracting phase was filtered through a
single-use 0.22µm nylon syringe filter (Aldrich) pass
through disposable filter 2.5um prior to analysis on
HPLC system under optimum separation condition,
the concentre of residual hormone were quantitatively
determined by comparison the resulted peak areas of
authentic standard with the peak area of samples under
the same separation condition.
Sample Injection
The list in (Table 2 represents the sequence of the
eluted material of the standard (1µg/mL).
Table 2. Sequence of the eluted material of the
standard (1µg/mL)
Hormones
Retention
time
Peak
area
Concentration
(µg/mL)
Estrone
0.553
42003
10
Zeranol
1.64
28684
10
Diethylstilbestrol
2.53
15531
10
Trembolone
3.89
14678
10
Medroxy
progesterone
4.70
17622
10
Melengestrole
acetate
5.64
19788
10
Progesterone
6.56
21383
10
Estradiol
benzoate
7.46
24298
10
* The residue concentrations from each injection
(10µl) obtained from the peaks calculations and
modified to µg/kg: [Meat sample] = peak area of
samples/peak area of standrd×1×10
* 1= [Meat sample] concentration of standard/µg/ml
modified to µg/kg (ppb)
* 10= dilution factor
Statistical Analysis
The data were calculated and statistically processed
using the statistical package SPSS 18.0 (SPSS,
Chicago, IL, USA). The level of statistical significance
was set at 0.05 for t-test and ANOVA.
RESULTS AND DISCUSSION
The extraction efficiency by liquid-liquid extraction
was directly related to the extraction solvent used for
tissue sample. As glacial acetic acid (0.1 mol/L), 0.1
mol/L ethanol, sodium acetate, dichloromethane and
hexane are the common for extracting steroids from
animal tissues were investigated to evaluate their
efficiency for extracting steroids from animal tissue.
The results showed that the liquid-liquid extraction is
advisable and applicable method of extraction and
high efficiency with less matrix interferences was
obtained from glacial acetic acid and ethanol
extraction system. Therefore, acid and alcohol were
employed as the extraction solvent in this work.

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Table 3. Hormone concentration (µg/kg) resulted in HPLC-UV chromatograms
*Kilcoy, *Pure South,*Veal Topsides, *P.Van Horten & *Zn and Creek Stone Farms, are the trade name of the meat
samples which also refer the name of meat produced companies which are *Australia, *Newzland, *Switzerland,
*Holand and *America respectively.
Figure 1.Chromatogram of high performance liquid chromatography-Ultra Violet (HPLC-UV) of standards and frozen
meat samples of Kilcoy solutions of hormonal residues by using Mobile phase (A) 0.1% Ammonium Formate in de-
ionized water. Solvents V/Methanol: Acetonitrile HPLC grade, and linear gradients0% B to 100%B. 1-Estrone, 2-
Zeranol, 3-Diethylstilbestrol, 4- Trembolone, 5-Medroxy progesterone, 6- Melengestrole acetate, 7- Progestrone,
Estradiol benzoate
Trade Name
Hormones
Estrone
Zeranol
Diethylstilbestrol
Trenbolone
Medroxyprogestrone
Melengestrol Acetate
Progestrone
Estradiol benzoate
*Kilcoy
*Pure
.South
*Veal
Topsides
*P.Van
Horten & Zn
*CreekStone
Farms
Australia
Newzland
Switzerland
Holand
America
0.36
0.309
0.476
0.373
0.318
0.483
0.4075
0.405
0.623
0.405
0.582
0.871
0.749
1.893
0.543
0.851
0.504
0.818
0.838
0.44
0.492
0.63
0.81
1.104
0.706
0.581
0.834
0.622
0.973
0.764
0.531
0.61
0.5015
0.855
0.742
0.645
0.368
0.289
0.425
0.364

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Figure 2. Chromatogrm of high performance liquid chromatography-Ulra Violet (HPLC-UV) of standards and frozen
meat sample of Pure South solutions for hormonal residues by using Mobile phase (A) 0.1% Ammonium Formate in
de-ionized water. Solvents V/Methanol: Acetonitrile HPLC grade, and linear gradients0% B to 100%B. 1-Estrone, 2-
Zeranol, 3-Diethylstilbestrol, 4- Trembolone, 5-Medroxy progesterone, 6- Melengestrole acetate, 7- Progestrone,
Estradiol benzoate
Figure 3. Chromatogram of high performance liquid chromatography- Ultra Violet (HPLC-UV) of standards and
frozen meat sample of Pure south solutions for hormonal residues by using Mobile phase(A) 0.1% Ammonium
Formate in de-ionized water. Solvents V/Methanol: Acetonitrile HPLC grade, and linear gradients0% B to 100%B. 1-
Estrone, 2- Zeranol, 3-Diethylstilbestrol, 4- Trembolone, 5-Medroxy progesterone, 6- Melengestrole acetate, 7-
Progestrone, Estradiol benzoate

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Figure 4. Chromatogram of high performance liquid chromatography-Ultra Violet (HPLC- UV) of standards and
frozen meat sample of P.Van Horten & Zn solutions of hormonal residues by using Mobile phase (A) 0.1%
Ammonium Formate in de-ionized water. Solvents V/Methanol: Acetonitrile HPLC grade, and linear gradients0% B
to 100% B. 1-Estrone, 2- Zeranol, 3-Diethylstilbestrol, 4- Trembolone, 5-Medroxy progesterone, 6- Melengestrole
acetate, 7- Progestrone, Estradiol benzoate
Figure 5. Chromatogram of high performance liquid chromatography-Ultra Violet (HPLC-UV) of standards and
frozen meat sample of Creek Stone Farms solutions of hormonal residues by using Mobile phase (A) 0.1% Ammonium
Formate in de-ionized water. Solvents V/Methanol: Acetonitrile HPLC grade, and linear gradients0% B to 100%B. 1-
Estrone, 2- Zeranol, 3-Diethylstilbestrol, 4- Trembolone, 5-Medroxy progesterone, 6- Melengestrole acetate, 7-
Progestrone, Estradiol benzoate.
Meat and meat products play an important role in
human nutrition; therefore, it should be safe and does
not contain factors or substances harmful for human
health (Clark and Henderson, 2003). However, the
anabolic agents used for various purposes in animal
husbandry for slaughter, could leave residues and
hence cause consumers health issue (Brusca et al.
2014). The European Economic Community (EEC)
banned the use of anabolic compounds as growth
accelerators in food animals, while the United States
Food and Drug Administration (USFDA) permitted
the limited use of some hormones with natural origins
(such as estradiol) and some synthetic hormones (such
as zeranol and trenbolone) (Nielen et al, 2007).
According to (WHO, 2006) beef muscle have to be
lack of estrone residues; whereas, the results in this
study presented the high values of estrone in which no
of the samples from the five countries of Australia,
Newzland, Switzerland, Holand and America were in

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Table 4. The evaluation of the average values of the anabolic residues (µg/ kg) of the meat samples from statistical aspects
Hormones
Trade Name
Estrone
Zeranol
Diethyl
stilbestrol
Trenbolone
Medroxy
progestrone
Melengestrol
Acetate
Progesterone
Estradiol
benzoate
Kilcoy
0.360 ±0.25
0.483±0.23
0.582ab±0.26
0.851±0.40
0.492a±0.28
0.581±0.18
0.531±0.34
0.645±0.34
Pure South
0.309±0.12
0.407±0.13
0.871ab±0.23
0.504±0.31
0.630ab±0.40
0.834±0.30
0.610±0.30
0.368±0.50
Veal Topsides
0.476±0.51
0.405±0.38
0.749ab±0.41
0.818±0.90
0.810ab±0.29
0.622±0.31
0.501±0.36
0.289±0.69
P.Van Horten &
Zn
0.373±0.13
0.623±0.26
1.893b±1.23
0.838±0.49
1.104b±0.47
0.973±0.38
0.855±0.37
0.425±0.48
Creek Stone
Farms
0.318±0.29
0.405±0.75
0.543a±0.43
0.440±0.25
0.706ab±0.27
0.764±0.21
0.742±0.45
0.364±0.49
Total
0.3672 ±0.26
0.4647 ±0.35
0.9276*±0.51
0.6902 ±0.47
0.7484*±0.34
0.7548±0.27
0.6479 ±0.36
0.4182± 0.50
a, b, c : The differences between the groups which have different letters under the same subgroup columns are statistically significant (P < 0.05).
* : The difference of hormone residue level in meat samples statistically important (p< 0.05).

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Table 5. Maximum hormones residue limits (MHRLs), present in beef muscles from non- treated and treated cattle (compilation of data reported by
(EFSA, 2007), (JECFA, 2005) and (Codex, 2007) which established for beef muscles).
*a: Hormone residue in Non-treated animals * b: Hormone residue in Treated animals with hormones.
A=Kilcoy B= Pure South C= Veal Topsides D= P.Van Horten & Zn E= Creek Stone Farms
Hormones substances
Inter-
national limits
Reference
[Hormones] resulted in HPLC-UV
A B C D E
Estrone
*a.0.0 pg/g
*b. 0
(WHO,2006)
0.360 ±0.259
0.309±0.126
0.476±0.517
0.373±0.138
0.318±0.298
Zeranol
*a. 0.01-0.73 ppb
*b. 0–0.5µg/kg
(FSIS,2002)
0.483±0.236
0.407±0.130
0.405±0.383
0.623±0.263
0.405±0.754
Diethylstilboestrol
*a.0.15 µg/kg
*b.<10µg/kg
(FDA 2013)
0.582±0.268
0.871±0.231
0.749±0.413
1.893±1.238
0.543±0.431
Trenbolone
*a. 0- 0.02 µg/kg
*b. 0.18-0.28 ppb
(JECFA/Codex,2007)
0.851±0.401
0.504±0.317
0.818±0.909
0.838±0.493
0.440±0.253
Medroxy progesterone
*a.0.23
*b.0.53
(Codex, 2007)
0.492±0.280
0.630±0.402
0.810±0.297
1.104±0.474
0.706±0.272
Melengestrol Acetate
*a. -
*b. 0–0.03 µg/kg
(FSIS,2002)
(FSIS,2002)
0.581±0.188
0.834±0.301
0.622±0.315
0.973±0.384
0.764±0.212
Progesterone
*a. 0.23- 0.77 ppb
*b. 3 µg/kg
*a. 0-30µg/kg
(JECFA/Codex,2007)
(FDA 2013)
0.531±0.342
0.610±0.309
0.501±0.367
0.855±0.379
0.742±0.457
Estradiol benzoate
*b. 5 ppb
(EFSA, 2007)
0.645±0.347
0.368±0.502
0.289±0.691
0.425±0.489
0.364±0.494

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safe level of estrone regarding the Kilcoy, Pure South,
Veal Topsides, P.Van Horten & Zn, Creek Stone
Farms companies (respectively) meat products (see
table 3 and 4). This indicates that acceptable tolerance
limits mentioned above should be immediately taken
into account for usage of estrone in animal husbandry
and its control in meat and meat products for
mentioned companies have to be focused.
Moreover, the use of zeranol was also banned by the
European Community in 1985 ( Jodlbauer et al.,
2000); whereas, very small (0–0.5µg/kg) amount
allowed to be available in treated animal in 2007
(EFSA, 2007). On the other hand, the zeranol level
was much higher in the samples of all the mentioned
five countries than the international limits and there
was no significant difference (P value= 0.56) between
the countries regarding zeranol level. This is also
indicated that the frozen beef samples of Kilcoy, Pure
South, Veal Topside, P.Van Horten & Zn and Creek
Stone Farms are not in allowed in terms of zeranol
value (see table 5). Furthermore, diethylstilbestrol
which is a carcinogenic and is not metabolized by the
organism, has been banned in animal husbandry and
not permitted to be present in food stuffs (EC,
1999).Consequently, the results presented that the
slaughtered cattle has been treated with
diethylstilboestrol and the residues in all the
companies samples were much more than the
international limits (<10µg/kg) (FDA, 2013).
Trenbolone was also detected in higher (0.18-0.28
ppb) in comparison with the national limits (0.69ppb)
(JECFA/Codex, 2007) in all the five groups of
samples. Medroxy progesterone and melengestrol
acetate levels were also not in safe level (0.7484ppb
and 0.7548ppb respectively) except in Kilcoy which
was (0.49ppb)( see table 4), the rest four countries
have extra containing medroxy progesterone and
melengestrol acetate in their products. According to
table 3, the cattle muscles of all the 5 trademarks were
treated with progesterone and estradiol benzoate;
whereas, the level of both hormones were (0.6479 and
0.4182 respectively) in safe level. Steroids can remain
parts per billion even 2 or 3 months after implantation;
understandably, it can be stated that the hormones
were higher than the international limits, have been
implanted before slaughtering closely (Nazil et al,
2005). In addition, the Post Hoc tests of ANOVA
resulted that there is no significant difference of each
estrone, zeranol trenbolone, melengestrol acetate,
progesterone and estradiol among the five trademarks
as the P values were, 0.649, 0.567, 0.40, 0.80, 0.26
and 0.29 respectively. There were also not significant
differences between Kilcoy, Pure South and Veal
Topsides regarding Diethylstilbestrol concentration,
but also the difference was only between P.Van Horten
& Zn and Creek Stone Farms(P value=0.046). In
addition, P.Van Horten & Zn again was significantly
different from Kilkoy in Medroxy progesterone level
(P value=0.03); whereas, no significant difference
seen in Medroxy progesterone concentration among
the rest companies. According to (Shao et al, 2005),
the conducting recovery tests before the study will be
useful for a correct test result. Understandably, the
results of this work are highlighted as they give some
information about the hormone residues and
comparing with some international standards to
improve the meat industry and animal husbandry.
CONCLUSION
This study presents that the anabolic steroids might be
available in almost samples with different levels
regarding each company and country, which indicates
the importance of analyzing the residue level of the
anabolic agents in different imported meat, since the
anabolic agents may pose a potential risk to public
health. Estrone, zeranol, diethylstilbestrol,
Trembolone, medroxy progesterone, melengestrole
acetate, progesterone, estradiol benzoate are still
implanting into cattle to promote growth rate;
however, progesterone, estradiol benzoate residues
were in safe level in this work. While the samples of
Kilcoy, Pure South, Veal Topsides, P.Van Horten &
Zn and Creek Stone Farms could be harmful and have
to be banned.
Acknowledgements
This paper is a result of the research within the project
MH5vet “Confirmation and Quantification of Eight
Anabolic Steroids in Imported Frozen Beef Muscles in
Sulaimani Markets Using HPLC-UV” which financed
by Theriogenology and Microbiology department-
College of Veterinary Medicine University of
Sulamiani-Kurdistan.
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